EGF receptor and other members of the ErbB family of receptor tyrosine kinases (RTKs) play essential physiological roles in development and maintenance of epithelial tissues by generating cell proliferation, survival, differentiation, and migration signals in response to specific ligands. Activation of ErbB receptors is also linked to the initiation and progression of human cancers. These RTKs have emerged as important therapeutic targets of anti-receptor antibodies and kinase inhibitors, although both therapeutic modalities suffer from rapid resistance development, necessitating alternate approaches to ErbB-targeted therapy. An essential pre-requisite for cellular response to EGFR and other RTK ligands is that an activation-competent pool of RTKs must be displayed on the cell surface. ErbB receptor overexpression increases this activation-ready pool. Sufficiently high levels of ErbB receptor overexpression induce ligand-independent activation, which can also be achieved by cancer-associated activating mutations. A fundamental feature of RTK function is that the newly synthesized as well as ligand-internalized receptors undergo a sorting process that determines whether they will recycle back to the cell surface for ligand binding and signaling or will be targeted for lysosomal degradation. We, and others, have shown that Cbl family of ubiquitin ligases are essential regulators of the lysosomal fate. Studies carried out by others and by us during the current funding period have led to a novel hypothesis that members of the C-terminal Eps15-homology (EH) domain-containing (EHD) protein family function as key regulators of the recycling fate of receptors. In this competing renewal application, we will employ unique EHD knockout mouse models and cellular reagents derived from these animals together with a vast array of immunological, biochemical and cellular reagents that have been generated to test the hypotheses: EHD protein-dependent endocytic recycling is a key controller of the cell surface display and recycling fate of EGFR; loss of EHD function will attenuate the ability of EGFR to propagate oncogenic signals in vitro and will abrogate EGFR-driven oncogenesis in vivo; and abrogation of endocytic recycling will enhance the efficacy of EGFR targeted therapy with an inhibitory antibody. Thus, this proposal will evaluate the endocytic recycling of EGFR as a novel therapeutic target in EGFR-driven cancer. Insights gained from these studies should further our understanding of the molecular and cell biological regulators of oncogenic signaling by RTKs, and help validate the endocytic recycling of RTKs as a new approach to rationally design anti-cancer agents to potentiate existing RTK-targeted therapies. PUBLIC HEALTH RELEVANCE: Abnormal activities of certain receptors on normal breast cells drive them to become cancerous. Our laboratory studies a group of genes whose products control the levels and activity of these receptors on our cells, such that these gene products become part of the network that drives a normal cell to become cancerous. Based on initial molecular work, further studies are proposed to investigate using cell culture and animal models to decipher the exact contributions of this new molecular mechanism in the context of breast cancer. By focusing on genetic strategies by which the molecules under question will be turned off to assess if cancer progression is slowed or halted, our studies will attempt to validate if the new molecular pathway could serve as a target for future drug development. As receptors of the type studied in this project contribute to breast, lung and other cancers, lessons learned here are hoped to open avenues for designing new drugs to treat human cancers and to aid in deciding the type of therapy a patient might receive thus sparing those unlikely to benefit from toxic therapies. While for practical reasons our studies will focus on breast cancer, the insights generated here could be helpful in most cancers as well as in other diseases where abnormal receptor function is involved, such as diabetes and autoimmunity.